Bottom Line:
NK cells were shown to be responsible for focal inflammation, and to be induced to migrate at high levels, in MCMV-infected livers.MIP-1alpha gene expression was elevated at coinciding times, and mice deficient in MIP-1alpha function were dramatically inhibited in both inflammatory and protective liver responses.The results precisely define MIP-1alpha-dependent steps required to achieve NK cell inflammation during, and mechanisms promoting defense against, viral infections in tissues.

Figure 5: Absence of liver inflammatory foci and decreased antiviral states in MCMV-infected MIP-1α–deficient mice. Liver samples were prepared from control C57BL/6 and C57BL/6–MIP-1α−/− day 2 MCMV-infected mice. Organs were harvested, and segments were either fixed for paraffin embedding, sectioning, and H&E staining, or frozen and then homogenized for viral titers in plaque assays as described in Materials and Methods. A and B show morphology of H&E-stained sections isolated from (A) C57BL/6 and (B) C57BL/6-MIP-1α−/− mice. Both A and B present areas encompassing a central vein (cv, central vein, given in A for orientation). Large arrow in A denotes inflammatory foci shown in inset. Large arrow in B denotes cluster of cytomegalic inclusion bodies shown in inset, and small arrows point to other clusters of intranuclear inclusion bodies radiating from the central vein. Photographs were taken at ×31.25. Inset photographs were taken at ×125. Bar, 100 μm. C gives inflammatory foci, D gives total numbers of nucleated cells, and E presents frequencies of cytomegalic inclusion bodies per defined area of 8 × 1 mm2 liver sections. C–E show mean results (± SE) from three replicate mice with samples harvested from infected mice on day 2 in hatched bars and on day 3 in solid bars. Differences between control C57BL/6 and C57BL/6-MIP-1α−/− mice are significant, * P <0.0005.

Mentions:
Studies were carried out to determine if MIP-1α deficiencies and an associated lack of NK cell–inflammatory responses altered early resistance to MCMV infection. Livers were isolated from day 2 infected C57BL/6 and C57BL/ 6-MIP-1α−/− mice, and prepared for either morphological analyses or determination of viral titers in plaque assays. Enumeration of inflammatory foci and total numbers of nucleated cells per defined liver area demonstrated that the absence of endogenous MIP-1α profoundly inhibited, i.e., >99%, inflammation in the liver (Fig. 5, A, B, C and D). Inflammatory responses were not simply delayed because inhibitory effects were maintained in samples taken on day 3 after infection (Fig. 5, C and D). In contrast, viral infection increased in the liver (Fig. 5 E). Cells having the morphological characteristics of MCMV infection, i.e., cytomegalic inclusion bodies, were infrequent in tissues from infected mice having MIP-1α (Fig. 5 A). They were increased by over fourfold in livers from infected C57BL/6-MIP-1α−/− as compared to C57BL/6 mice (Fig. 5, B and E). Consistent with our previous studies of NK cell–depleted mice (18), there were no differences in liver viral titers between C57BL/6 control and MIP-1α−/− mice on day 2 after infection, but there were significant rises in liver viral titers in C57BL/6-MIP-1α−/− mice on day 3. At this time, mean log PFU per gram of liver ± SE was 4.8 ± 0.3 in C57BL/ 6 mice and 5.9 ± 0.3 in C57BL/6-MIP-1α−/− mice (P <0.01). Thus, absence of MIP-1α results in decreased inflammation but increased susceptibility to MCMV infection in the liver.

Figure 5: Absence of liver inflammatory foci and decreased antiviral states in MCMV-infected MIP-1α–deficient mice. Liver samples were prepared from control C57BL/6 and C57BL/6–MIP-1α−/− day 2 MCMV-infected mice. Organs were harvested, and segments were either fixed for paraffin embedding, sectioning, and H&E staining, or frozen and then homogenized for viral titers in plaque assays as described in Materials and Methods. A and B show morphology of H&E-stained sections isolated from (A) C57BL/6 and (B) C57BL/6-MIP-1α−/− mice. Both A and B present areas encompassing a central vein (cv, central vein, given in A for orientation). Large arrow in A denotes inflammatory foci shown in inset. Large arrow in B denotes cluster of cytomegalic inclusion bodies shown in inset, and small arrows point to other clusters of intranuclear inclusion bodies radiating from the central vein. Photographs were taken at ×31.25. Inset photographs were taken at ×125. Bar, 100 μm. C gives inflammatory foci, D gives total numbers of nucleated cells, and E presents frequencies of cytomegalic inclusion bodies per defined area of 8 × 1 mm2 liver sections. C–E show mean results (± SE) from three replicate mice with samples harvested from infected mice on day 2 in hatched bars and on day 3 in solid bars. Differences between control C57BL/6 and C57BL/6-MIP-1α−/− mice are significant, * P <0.0005.

Mentions:
Studies were carried out to determine if MIP-1α deficiencies and an associated lack of NK cell–inflammatory responses altered early resistance to MCMV infection. Livers were isolated from day 2 infected C57BL/6 and C57BL/ 6-MIP-1α−/− mice, and prepared for either morphological analyses or determination of viral titers in plaque assays. Enumeration of inflammatory foci and total numbers of nucleated cells per defined liver area demonstrated that the absence of endogenous MIP-1α profoundly inhibited, i.e., >99%, inflammation in the liver (Fig. 5, A, B, C and D). Inflammatory responses were not simply delayed because inhibitory effects were maintained in samples taken on day 3 after infection (Fig. 5, C and D). In contrast, viral infection increased in the liver (Fig. 5 E). Cells having the morphological characteristics of MCMV infection, i.e., cytomegalic inclusion bodies, were infrequent in tissues from infected mice having MIP-1α (Fig. 5 A). They were increased by over fourfold in livers from infected C57BL/6-MIP-1α−/− as compared to C57BL/6 mice (Fig. 5, B and E). Consistent with our previous studies of NK cell–depleted mice (18), there were no differences in liver viral titers between C57BL/6 control and MIP-1α−/− mice on day 2 after infection, but there were significant rises in liver viral titers in C57BL/6-MIP-1α−/− mice on day 3. At this time, mean log PFU per gram of liver ± SE was 4.8 ± 0.3 in C57BL/ 6 mice and 5.9 ± 0.3 in C57BL/6-MIP-1α−/− mice (P <0.01). Thus, absence of MIP-1α results in decreased inflammation but increased susceptibility to MCMV infection in the liver.

Bottom Line:
NK cells were shown to be responsible for focal inflammation, and to be induced to migrate at high levels, in MCMV-infected livers.MIP-1alpha gene expression was elevated at coinciding times, and mice deficient in MIP-1alpha function were dramatically inhibited in both inflammatory and protective liver responses.The results precisely define MIP-1alpha-dependent steps required to achieve NK cell inflammation during, and mechanisms promoting defense against, viral infections in tissues.